Controlling AR spread: a novel approach

AR has represented a serious impediment to antibiotic therapy for as long as antibiotics have been used. Although mutations are responsible for some specific cases of AR, the driving force behind the problem of multiresistance to antimicrobials is gene acquisition by human pathogens. Understanding and control of AR spread was largely through a unique approach: the precise description of AR genes presently found in hospitals, and inference from this of the working mechanisms of dispersion. This approach resulted in the accumulation of an impressive knowledge base of individual elements, modules and clones of bacteria that underpin AR dissemination. Nevertheless, and in spite of the vast number of publications on the subject, after over 40 years of study, we are as yet unable to circumvent or even simply restrain AR dissemination.

We believe that a novel approach is required in order to facilitate a decisive step forward. This approach will necessitate an understanding of the molecular mechanisms sustaining the observed behaviour of mobile genetic elements (MGEs) and the forces driving the evolution of AR genes themselves, their recruitment from environmental bacteria, their dissemination by dispersion modules and their final acquisition (establishment and stable inheritance) by bacterial pathogens. It will be important to answer essential questions such as the origin of AR genes, how they are mobilized from their original locations by different MGEs (integrons, plasmids and transposons), why some bacterial hosts readily acquire and transmit some elements but not others and whether the underlying mechanisms impose difficulties and bottlenecks in transmission. Answers to these questions will help us to better evaluate the present situation, to assess the overall dynamics of AR transmission, to diagnose the immediate future and to suggest novel strategies for the control of AR dissemination. This will be particularly important in the hospital environment, where the problem of multiresistance dissemination is painfully acute.

The challenge: understanding mechanisms and process dynamics

Although the main avenues for AR dissemination are fairly well known (see the state of the art description (section 4.1)), we still lack both a deep knowledge of many of the detailed mechanisms involved as well as of the influence that different factors (genetic and environmental) have on the modes of AR dissemination and final acquisition by pathogens. In a few cases where a more intimate knowledge of these mechanisms is available, for instance, the mechanisms of plasmid stability, this knowledge is already offering new tools for biotechnology. In summary, we propose to investigate on the mechanisms of AR transmission in the hope of identifying and characterizing targets for novel biotechnological intervention.

Thus, we will endeavour to achieve three main goals in the CRAB project:

1. To gain a deeper fundamental knowledge of the main mechanisms of AR dissemination

2. To ascertain the influences of the genetic bacterial background and the human environment in the processes of dissemination

3. To apply the knowledge we already have of one of these processes (plasmid stability) for the construction of new tools for biotechnology.

The precise objectives of CRAB deal with each of the dissemination modules acting in the chain of transfer from the environmental source to the ultimate host, the bacterial pathogen– integrons (Work package (WP) 1), transposons (WP2), conjugative plasmids (WP3) and stability modules (WP4) – in a concerted approach.

We concentrate on the as yet improperly studied aspects of these mechanisms with the following objectives:

WP1 : Establishing the mechanisms of integron cassette recombination, identification of the signals triggering integrase expression and the molecular evolution of gene cassettes.

WP2 : Establishing the propensities of three major classes of ISs to acquire, stabilise and vehicle AR genes and identification of host factors that affect transposition.

WP3 : Identification of host factors that affect conjugation, development of in vitro models for conjugation on cultured mammalian cells.

WP4 : Identification of novel stabilisation systems and their intracellular targets and identification of host factors that trigger these systems

Each work package will tackle the three main goals stated above, but taking into account the specifics of each system.